Preparation and In Vitro Evaluation of CD3Fab-Decorated Lipoplexes to Improve Delivery of siRNA Targeting C1858T PTPN22 Variant in T Lymphocytes for Type 1 Diabetes immunotherapy.

Type 1 diabetes (T1D) and autoimmune thyroid diseases are the most common autoimmune endocrinopathies. Nowadays, substitutive hormonal administration is the only treatment. As regard T1D, insulin does not fully replace the body's insulin requirements and T1D remains associated with acute and chronic complications. This emphasizes the need to identify personalized effective immunotherapeutic strategies targeting T and B lymphocytes as important players in the autoimmune response. Recently, particular interest was generated by the potential pathophysiological role of the C1858T variant in the protein tyrosine phosphatase non-receptor type N22 (PTPN22) gene, which changes amino acid residue 620 from Arg to Trp (R620W) in the lymphoid tyrosine phosphatase Lyp. The R620W variant affects both the innate and adaptive immunity leading to a gain of function mutation with paradoxical reduced T cell activation. We previously exploited a novel personalized immunotherapeutic approach based on siRNA delivered by liposomes (lipoplexes) that selectively inhibit variant allele (PCT/IT2019/050095). The goal of the present study is improving lipoplexes carrying siRNA for variant C1858T to enhance delivery to B and T cells by functionalizing them with Fab of MoAb. Rituximab (Rituxi)Fab-Lipoplexes specifically bind to B cells and have impact on their function in T1D PBMC upon CpG stimulation showing a higher inhibitory effect on total cell proliferation and IgM+ plasma cell differentiation compared to the not functionalized ones (PMID35008834). Similarly, CD3Fab-Lipoplexes specifically bind T lymphocytes of the Jurkat human T lymphoblastoid cell line and within PBMC of healthy donors with a higher percentage of binding compared to non-functionalized lipoplexes, suggesting that functionalization does not affect CD3Fab biofunctionality. These results open the pathway to functionalized lipoplexes with FDA approved MoAb i.e. Teplixumab/Otelixizumab for specific targeting of T cells in T1D.